R03 Resubmission Role of Sox4 in B cell development Xiaoping Sun, M.D., Ph.D. B cells constitute an integral part of the immune system. B cell deficiency or malfunction can result in impaired humoral immunity and is one of the most common causes for primary immunodeficiency diseases (PIDs). Of more than 120 known PIDs, B cell deficiencies and combined B and T cell deficiencies are responsible for nearly 40. B cell development is a sequential process including the following stages: hematopoietic stem cell, multilineage progenitor, common lymphoid progenitor, pre-pro-B, pro-B, pre-B, immature B, and mature B. The process is regulated by a number of hierarchically ordered genes, especially those encoding transcription factors. However, it is not well understood how these critical genes integrate signals and regulate this highly ordered developmental process. The Sox4 gene, which encodes a transcription factor, is one such gene. Sox4 knockout was shown 13 years ago to cause B cell developmental arrest, indicating that the function of Sox4 is indispensable for this process. However, no further study has been reported on the role of Sox4 in B cell development. This is in part due to the fact that conventional Sox4-knockout is embryonically lethal and the experimental tools that are widely used today have only recently become available. By using a conditional knockout mouse strain, we have characterized the effect of Sox4 deficiency on B cell development in adult mice. We found that pre-pro-B cells are slightly decreased in number, whereas pro-B cells and later stage B cells are nearly absent when Sox4 is knocked out early in development. However, we found no significant effect when Sox4 is knocked out in mature B cells. On the basis of these findings, we have formulated our central hypothesis: it is in the pre-pro-B cells that Sox4 is functioning to promote the transition from pre-pro-B to pro-B stage. We will test this hypothesis by restoring Sox4 function specifically in pre-pro-B cells to see whether such restoration rescues subsequent B cell development. We will also rule out the possibility that the B cell developmental arrest is owing to the inability of pro-B cells to proliferate and/or increased pro-B cell apoptosis. Moreover, we will test whether any of a group of genes known to be essential for B cell development is regulated by Sox4. In addition, ChIP-on-ChIP and gene expression microarray assays will be performed to systematically identify and characterize Sox4 downstream genes. Successful completion of this research will reveal the mechanism(s) by which Sox4 affects B cell development and the genes that are regulated by Sox4. We expect that this research, by paving the way for further characterization of Sox4 in early B cell development, will advance our understanding of this process, where dysregulation may cause immunodeficiency, autoimmune disease, and B cell malignancy. PUBLIC HEALTH RELEVANCE: R03 Resubmission Role of Sox4 in B cell development Xiaoping Sun, MD, PhD B cell deficiency, as one of the major causes of PIDs, results in deficiency in immunoglobulins and hence impairs humoral immunity. The goal of the proposed research is to reveal the molecular and cellular mechanism underlying the function of Sox4, a protein whose deficiency disrupts B cell development. Successful completion of this proposed research will help us better understand PIDs and develop preventive and therapeutical measurements against PIDs.